Maternal IL-33 critically regulates tissue remodeling and type 2 immune responses in the uterus during early pregnancy in mice.

The pregnant uterus is an immunologically rich organ, with dynamic changes in the inflammatory milieu and immune cell function underlying key stages of pregnancy. Recent studies have implicated dysregulated expression of the interleukin-1 (IL-1) family cytokine, IL-33, and its receptor, ST2, in poor pregnancy outcomes in women, including recurrent pregnancy loss, preeclampsia, and preterm labor. How IL-33 supports pregnancy progression in vivo is not well understood. Here, we demonstrate that maternal IL-33 signaling critically regulates uterine tissue remodeling and immune cell function during early pregnancy in mice. IL-33-deficient dams exhibit defects in implantation chamber formation and decidualization, and abnormal vascular remodeling during early pregnancy. These defects coincide with delays in early embryogenesis, increased resorptions, and impaired fetal and placental growth by late pregnancy. At a cellular level, myometrial fibroblasts, and decidual endothelial and stromal cells, are the main IL-33+ cell types in the uterus during decidualization and early placentation, whereas ST2 is expressed by uterine immune populations associated with type 2 immune responses, including ILC2s, Tregs, CD4+ T cells, M2- and cDC2-like myeloid cells, and mast cells. Early pregnancy defects in IL-33-deficient dams are associated with impaired type 2 cytokine responses by uterine lymphocytes and fewer Arginase-1+ macrophages in the uterine microenvironment. Collectively, our data highlight a regulatory network, involving crosstalk between IL-33-producing nonimmune cells and ST2+ immune cells at the maternal-fetal interface, that critically supports pregnancy progression in mice. This work has the potential to advance our understanding of how IL-33 signaling may support optimal pregnancy outcomes in women.

Decidualization modulates the mesenchymal stromal/stem cell and pericyte characteristics of human decidual stromal cells. Effects on antigen expression, chemotactic activity on monocytes and antitumoral activity.

Decidual stromal cells (DSCs) are the most abundant cellular component of human decidua and play a central role in maternal-fetal immune tolerance. Antigen phenotyping and functional studies recently confirmed the relationship of DSCs with mesenchymal stem/stromal cells (MSCs) and pericytes, the latter two cell types being closely related or identical. The present study investigated the effect of decidualization, a process of cell differentiation driven by progesterone (P4) and other pregnancy hormones, on the MSC/pericyte characteristics of DSCs. To this end we isolated undifferentiated DSC (preDSC) lines that were decidualized in vitro (dDSC) by the effect of P4 and cAMP. Using flow cytometry, we found significant downmodulation of the expression of the MSC/pericyte markers α-smooth muscle actin, nestin, CD140b, CD146 and SUSD2 in dDSCs. The dDSCs did not differ, compared to preDSCs, in the expression of angiogenic factors (characteristic of pericytes) HGF, FGF2, ANGPT1 or VEGF according to RT-PCR results, but had significantly increased PGF expression. In migration assays, preDSC-conditioned media had a chemotactic effect on the THP-1 monocytic line (characteristic of pericytes), and this effect was significantly greater in dDSC-conditioned media. Media conditioned with dDSC, but not with preDSC, induced apoptosis in 4 out of 6 different tumor cell lines (characteristic of MSCs) according to propidium iodide staining and flow cytometry results. Our findings show that decidualization induces phenotypic and functional changes in the MSC/pericyte properties of DSCs that may have a role in the normal development of pregnancy.

Single-cell transcriptomic atlas of the human endometrium during the menstrual cycle.

In a human menstrual cycle the endometrium undergoes remodeling, shedding and regeneration, all of which are driven by substantial gene expression changes in the underlying cellular hierarchy. Despite its importance in human fertility and regenerative biology, our understanding of this unique type of tissue homeostasis remains rudimentary. We characterized the transcriptomic transformation of human endometrium at single-cell resolution across the menstrual cycle, resolving cellular heterogeneity in multiple dimensions. We profiled the behavior of seven endometrial cell types, including a previously uncharacterized ciliated cell type, during four major phases of endometrial transformation, and found characteristic signatures for each cell type and phase. We discovered that the human window of implantation opens with an abrupt and discontinuous transcriptomic activation in the epithelia, accompanied with a widespread decidualization feature in the stromal fibroblasts. Our study provides a high-resolution molecular and cellular characterization of human endometrial transformation across the menstrual cycle, providing insights into this essential physiological process.

Pregnancy success in mice requires appropriate cannabinoid receptor signaling for primary decidua formation.

With implantation, mouse stromal cells begin to transform into epithelial-like cells surrounding the implantation chamber forming an avascular zone called the primary decidual zone (PDZ). In the mouse, the PDZ forms a transient, size-dependent permeable barrier to protect the embryo from maternal circulating harmful agents. The process of decidualization is critical for pregnancy maintenance in mice and humans. Mice deficient in cannabinoid receptors, CB1 and CB2, show compromised PDZ with dysregulated angiogenic factors, resulting in the retention of blood vessels and macrophages. This phenotype is replicated in Cnr1-/- but not in Cnr2-/-mice. In vitro decidualization models suggest that Cnr1 levels substantially increase in mouse and human decidualizing stromal cells, and that neutralization of CB1 signaling suppresses decidualization and misregulates angiogenic factors. Taken together, we propose that implantation quality depends on appropriate angiogenic events driven by the integration of CB2 in endothelial cells and CB1 in decidual cells.

Human PZP and common marmoset A2ML1 as pregnancy related proteins.

While pregnancy-related proteins (PRP) are known to contribute to immunotolerance during pregnancy, their significance to development of invasive placenta is unclear. We compared PRP expression in humans and the common marmoset (Callithrix jacchus), a new-world monkey. Invasive placenta was observed at the maternal-foetal interface of marmoset placenta from green fluorescent protein (GFP)-expressing foetus and wild type mother. The pregnancy zone protein (PZP) and alpha-2 macroglobulin-like 1 (A2ML1) proteins exhibited the most prominent increase in expression during the second trimester in humans and marmoset, respectively. In humans, PZP accumulated at the maternal-foetal interface and A2ML1 accumulated in the amnion. Similarly, A2ML1 mRNA was detected in marmoset placenta. These proteins belong to the A2M family of protease inhibitors, and both PZP and A2ML1 share around 90% homology between human and marmoset and have highly conserved structures. However, the protease-reacting bait regions of the proteins had lower homology (56.8-60.7% in proteins) relative to the rest of the sequence. Notably, the cleavage site of a proinflammatory proline-endopeptidase was preserved in human PZP and marmoset A2ML1. These proteins contain multiple sites that are cleaved by proteases involving proline-endopeptidase. Systemic regulation of these A2M family proteins may be important in animals with invasive placenta.

Distinctive phenotypes and functions of innate lymphoid cells in human decidua during early pregnancy.

During early pregnancy, decidual innate lymphoid cells (dILCs) interact with surrounding maternal cells and invading fetal extravillous trophoblasts (EVT). Here, using mass cytometry, we characterise five main dILC subsets: decidual NK cells (dNK)1-3, ILC3s and proliferating NK cells. Following stimulation, dNK2 and dNK3 produce more chemokines than dNK1 including XCL1 which can act on both maternal dendritic cells and fetal EVT. In contrast, dNK1 express receptors including Killer-cell Immunoglobulin-like Receptors (KIR), indicating they respond to HLA class I ligands on EVT. Decidual NK have distinctive organisation and content of granules compared with peripheral blood NK cells. Acquisition of KIR correlates with higher granzyme B levels and increased chemokine production in response to KIR activation, suggesting a link between increased granule content and dNK1 responsiveness. Our analysis shows that dILCs are unique and provide specialised functions dedicated to achieving placental development and successful reproduction.

Microgravity inhibits decidualization via decreasing Akt activity and FOXO3a expression in human endometrial stromal cells.

Decidualization is characterized by the differentiation of endometrial stromal cells (eSCs), which is critical for embryo implantation and maintenance of pregnancy. In the present study, we investigated the possible effect of simulated microgravity (SM) on the process of proliferation and in vitro decidualization using primary human eSCs. Exposure to SM for 36 h decreased the proliferation and migration of eSCs significantly, without inducing cell death and changes in cell cycle progression. The phosphorylation of Akt decreased under SM conditions in human eSCs, accompanied by a simultaneous decrease in the level of matrix metalloproteinase (MMP)-2 and FOXO3a. Treatment with Akti, an Akt inhibitor, decreased MMP-2 expression, but not FOXO3a expression. The decreased level of FOXO3a under SM conditions impeded autophagic flux by reducing the levels of autophagy-related genes. In addition, pre-exposure of eSCs to SM significantly inhibited 8-Br-cAMP induced decidualization, whereas restoration of the growth status under SM conditions by removing 8-Br-cAMP remained unchanged. Treatment of human eSCs with SC-79, an Akt activator, restored the reduced migration of eSCs and decidualization under SM conditions. In conclusion, exposure to SM inhibited decidualization in eSCs by decreasing proliferation and migration through Akt/MMP and FOXO3a/autophagic flux.

CSDC2, a cold shock domain RNA-binding protein in decidualization.

RNA-binding proteins (RBPs) have been described for cancer cell progression and differentiation, although there is still much to learn about their mechanisms. Here, using in vivo decidualization as a model, we describe the role of RBP cold shock domain containing C2 (CSDC2) in the endometrium. Csdc2 messenger RNA expression was differentially regulated depending on time and areas of decidua development, with the most variation in antimesometrium (AM) and, to a lesser degree, in the junctional zone (JZ). Immunohistochemistry of CSDC2 showed a preferentially cytoplasmic localization at AM and JZ, and nuclear localization in underneath myometrium and mesometrium (M). Cytoplasmic localization coincided with differentiated, DESMIN-marked areas, while nuclear localization coincides with proliferative zones. Uterine suppression of CSDC2 through intrauterine-injected-specific small interfering RNA (siRNA) led to abnormal decidualization in early pregnancy, with more extended antimesometrial area and with poor M development if compared with control siRNA-injected animals. These results suggest that CSDC2 could be a regulator during decidua development.

Pik3ca is required for mouse uterine gland development and pregnancy.

The PI3K/AKT signaling pathway plays a critical role in the maintenance of equilibrium between cell survival and apoptosis. The Pik3ca gene is mutated in a range of human cancers. It has been found to be oncogenic, and mutations lead to constitutive activation of the PI3K/AKT pathway. The expression patterns of PIK3CA proteins in the uterus of mice during early pregnancy indicate that it may play a role in the regulation of glandular epithelial cells, which is required to support uterine receptivity. To further investigate the role of Pik3ca in uterine function, Pik3ca was conditionally ablated only in the PGR-positive cells (Pgrcre/+Pik3caf/f; Pik3cad/d). A defect of uterine gland development and decidualization led to subfertility observed in Pik3cad/d mice. Pik3cad/d mice showed significantly decreased uterine weight compared to Pik3caf/f mice. Interestingly, a significant decrease of gland numbers were detected in Pik3cad/d mice compared to control mice. In addition, we found a decrease of Foxa2 expression, which is a known uterine gland marker in Pik3cad/d mice. Furthermore, the excessive proliferation of endometrial epithelial cells was observed in Pik3cad/d mice. Our studies suggest that Pik3ca has a critical role in uterine gland development and female fertility.

NLRP7 contributes to in vitro decidualization of endometrial stromal cells.

BACKGROUND: Nucleotide-binding oligomerization domain (NACHT), leucine rich repeat (LRR) and pyrin domain (PYD) 7 containing protein, NLRP7, is a member of the NLR family which serves as innate immune sensors. Mutations and genetic variants of NLRP7 have been found in women with infertility associated conditions, such as recurrent hydatidiform mole, recurrent miscarriage, and preeclampsia. Decidualization of endometrial stromal cells is a hallmark of tissue remodeling to support embryo implantation and proper placental development. Given defective decidualization has been implicated in miscarriage as well as preeclampsia, we aimed to explore the link between the NLRP7 gene and decidualization. METHODS: Endometrial samples obtained from pregnant women in the first trimester and non-pregnant women were used to study NLRP7 expression pattern. The human telomerase reverse transcriptase (hTERT)-immortalized human endometrial stromal cells (T-HESCs) were used to study the effect of NLRP7 on decidualization. Decidualization of T-HESCs was induced with 1 µM medroxyprogesterone acetate (MPA) and 0.5 mM 8-bromoadenosine 3':5'-cyclic monophosphate (8-Br-cAMP). siRNA was used to knock down NLRP7 while lentiviral vectors were used to overexpress NLRP7 in cells. NLRP7 expression was detected by immunofluorescence, qRT-PCR, and Western blotting. Decidualization markers, Insulin-like growth factor-binding protein 1 (IGFBP-1) and prolactin (PRL), were detected by qRT-PCR and ELISA. Nuclear translocation of NLRP7 was detected by the subcellular fractionation and confocal microscopy. The effect of NLRP7 on progesterone receptor (PR) activity was evaluated by a reporter system. RESULTS: NLRP7 was up-regulated in the decidual stromal cells of human first-trimester endometrium. After in vitro decidualization, T-HESCs presented with the swollen phenotype and increased expressions of IGFBP-1 and PRL. Knockdown or over-expression of NLRP7 reduced or enhanced the decidualization, respectively, according to the expression level of IGFBP-1. NLRP7 was found to translocate in the nucleus of decidualized T-HESCs and able to promote PR activity. CONCLUSIONS: NLRP7 was upregulated and translocated to the nucleus of the endometrial stromal cells in an in vitro decidualization model. Overexpressed NLRP7 promoted the IGFBP-1 expression and PR reporter activation. IGFBP-1 expression decreased with the knockdown of NLRP7. Therefore, we suggest that NLRP7 contributes to in vitro decidualization of endometrial stromal cells.

Autotaxin-lysophosphatidic acid-LPA3 signaling at the embryo-epithelial boundary controls decidualization pathways.

During pregnancy, up-regulation of heparin-binding (HB-) EGF and cyclooxygenase-2 (COX-2) in the uterine epithelium contributes to decidualization, a series of uterine morphological changes required for placental formation and fetal development. Here, we report a key role for the lipid mediator lysophosphatidic acid (LPA) in decidualization, acting through its G-protein-coupled receptor LPA3 in the uterine epithelium. Knockout of Lpar3 or inhibition of the LPA-producing enzyme autotaxin (ATX) in pregnant mice leads to HB-EGF and COX-2 down-regulation near embryos and attenuates decidual reactions. Conversely, selective pharmacological activation of LPA3 induces decidualization via up-regulation of HB-EGF and COX-2. ATX and its substrate lysophosphatidylcholine can be detected in the uterine epithelium and in pre-implantation-stage embryos, respectively. Our results indicate that ATX-LPA-LPA3 signaling at the embryo-epithelial boundary induces decidualization via the canonical HB-EGF and COX-2 pathways.

Role of selectins and their ligands in human implantation stage.

Selectins are a family of calcium-dependent, type I transmembrane, carbohydrate-binding glycoproteins. Selectins and their ligands are not only involved in physiological processes such as leukocyte homing and pathological processes such as cancer, but also play an essential role in the human implantation. L-selectin and its ligands participate in the adhesion of the blastocyst to the endometrium at the maternal-fetal interface. P-selectin and E-selectin are involved in immune recognition of maternal decidua to the embedded embryo as well as trophoblast migration within decidual spiral arterioles. Moreover, altered expression of selectins and their ligands are found to be associated with some abnormal pregnancies and infertilities. This review focuses on the current progress of research on the role of selectins and their ligands in the human implantation process.

Exposure to benzo[a]pyrene impairs decidualization and decidual angiogenesis in mice during early pregnancy.

Benzo[a]pyrene (BaP) is a ubiquitous environmental persistent organic pollutant and a well-known endocrine disruptor. BaP exposure could alter the steroid balance in females. Endometrium decidualization and decidual angiogenesis are critical events for embryo implantation and pregnancy maintenance during early pregnancy and are modulated by steroids. However, the effect of BaP on decidualization is not clear. This study aimed to explore the effects of BaP on decidualization and decidual angiogenesis in pregnant mice. The result showed that the uteri in the BaP-treated groups were smaller and exhibited an uneven size compared with those in the control group. Artificial decidualization was detected in the uteri of the controls, but weakened decidualization response was observed in the BaP-treated groups. BaP significantly reduced the levels of estradiol, progesterone, and their cognate receptors ER and PR, respectively. The expression of several decidualization-related factors, including FOXO1, HoxA10, and BMP2, were altered after BaP treatment. BaP reduced the expression of cluster designation 34 (CD34), which indicated that the decidual angiogenesis was inhibited by BaP treatment. In addition, BaP induced the downregulation of vascular endothelial growth factor A. These data suggest that oral BaP ingestion compromised decidualization and decidual angiogenesis. Our results provide experimental data for the maternal reproductive toxicity of BaP during early pregnancy, which is very important for a comprehensive risk assessment of BaP on human reproductive health.

Effects of neonatal exposure to a glyphosate-based herbicide on female rat reproduction.

In this study, we investigated whether neonatal exposure to a glyphosate-based herbicide (GBH) alters the reproductive performance and the molecular mechanisms involved in the decidualization process in adult rats. Newborn female rats received vehicle or 2 mg/kg/day of a GBH on postnatal days (PND) 1, 3, 5 and 7. On PND90, the rats were mated to evaluate (i) the reproductive performance on gestational day (GD) 19 and (ii) the ovarian steroid levels, uterine morphology, endometrial cell proliferation, apoptosis and cell cycle regulators, and endocrine pathways that regulate uterine decidualization (steroid receptors/COUP-TFII/Bmp2/Hoxa10) at the implantation sites (IS) on GD9. The GBH-exposed group showed a significant increase in the number of resorption sites on GD19, associated with an altered decidualization response. In fact, on GD9, the GBH-treated rats showed morphological changes at the IS, associated with a decreased expression of estrogen and progesterone receptors, a downregulation of COUP-TFII (Nr2f2) and Bmp2 mRNA and an increased expression of HOXA10 and the proliferation marker Ki67(Mki67) at the IS. We concluded that alterations in endometrial decidualization might be the mechanism of GBH-induced post-implantation embryo loss.

Uterine Epithelial Estrogen Receptor-α Controls Decidualization via a Paracrine Mechanism.

Steroid hormone-regulated differentiation of uterine stromal cells, known as decidualization, is essential for embryo implantation. The role of the estrogen receptor-α (ESR1) during this differentiation process is unclear. Development of conditional Esr1-null mice showed that deletion of this gene in both epithelial and stromal compartments of the uterus leads to a complete blockade of decidualization, indicating a critical role of ESR1 during this process. To further elucidate the cell type-specific function of ESR1 in the uterus, we created WE(d/d) mice in which Esr1 is ablated in uterine luminal and glandular epithelia but is retained in the stroma. Uteri of WE(d/d) mice failed to undergo decidualization, indicating that epithelial ESR1 contributes to stromal differentiation via a paracrine mechanism. We noted markedly reduced production of the leukemia inhibitory factor (LIF) in WE(d/d) uteri. Supplementation with LIF restored decidualization in WE(d/d) mice. Our study indicated that LIF acts synergistically with progesterone to induce the expression of Indian hedgehog (IHH) in uterine epithelium and its receptor patched homolog 1 in the stroma. IHH then induces the expression of chicken ovalbumin upstream promoter-transcription factor II, a transcription factor that promotes stromal differentiation. To address the mechanism by which LIF induces IHH expression, we used mice lacking uterine epithelial signal transducer and activator of transcription 3, a well-known mediator of LIF signaling. Our study revealed that LIF-mediated induction of IHH occurs without the activation of epithelial signal transducer and activator of transcription 3 but uses an alternate pathway involving the activation of the ERK1/2 kinase. Collectively our results provide unique insights into the paracrine mechanisms by which ESR1 directs epithelial-stromal dialogue during pregnancy establishment.

IMMUNOLOGICAL IMPLICATIONS IN MAMMALIAN SEMI-ALLOGENEIC PREGNANCY.

The mammalian conceptus is a semi-allograft. The maternal tolerance towards semi-allograft is primed by a battery of cytokines and resident decidual cells. Invasion of embryonic trophoblast, decidual vascular remodeling and unresponsive incipient 'Triple-negative' (CD3-, CD4- and CD8-) T-cells of foetus promote normal pregnancy. On the other hand, the maternal cytokine profile shift towards Th1 response is shown to enhance the risk during pregnancy. The periodical examination of maternal serum levels of Th1 and Th2 response mediated cytokines constitutes prognostic biomarkers to initiate therapeutic regimens.

[Effect of bone marrow cells transplantation on the decidua formation in pseudopregnant rats].

One of the most common causes of the current pregnancy loss is the failure of the decidual reaction of endometrial cells. It is assumed that a partial source of decidual cells in endometrial tissue is bone marrow cells (BMCs). In the present work, we have studied possible effect of BMCs transplantation on the process of decidualization using the model of pseudopregnancy in rats. BMCs were flushed from the rat femurs and tibias. The obtained suspension of single BMCs was injected into one of rat uterine horns on the 5th day of pseudopregnancy. PBS without cells was injected into the contralateral horn served as the control. Rats were sacrificed on the 11th day of pseudopregnancy. Decidua formed in the experimental uterine horn showed an increase in the meso-antimezometral direction of their diameter of about 1.5-2 times as compared with a control horn. The weight of decidual tissue in the experimental horn exceeded 3 times the weight of the control one. The presence of transplanted BMCs in decidual tissue was documented by preliminary double staining of BMCs with membrane dye PKH 26 Red and nuclear dye Hoechst 33342. Histological analysis of decidua sections after transplantation revealed any alterations neither in cell differentiation nor in tissue structure. We conclude that BMCs transplantation stimulates decidualization in animals.

Human decidual stromal cells secrete soluble pro-apoptotic factors during decidualization in a cAMP-dependent manner.

STUDY QUESTION: Is there a relationship between decidualization and apoptosis of decidual stromal cells (DSC)? SUMMARY ANSWER: Decidualization triggers the secretion of soluble factors that induce apoptosis in DSC. WHAT IS KNOWN ALREADY: The differentiation and apoptosis of DSC during decidualization of the receptive decidua are crucial processes for the controlled invasion of trophoblasts in normal pregnancy. Most DSC regress in a time-dependent manner, and their removal is important to provide space for the embryo to grow. However, the mechanism that controls DSC death is poorly understood. STUDY DESIGN, SIZE, DURATION: The apoptotic response of DSC was analyzed after exposure to different exogenous agents and during decidualization. The apoptotic potential of decidualized DSC supernatants and prolactin (PRL) was also evaluated. PARTICIPANTS/MATERIALS, SETTING, METHODS: DSC lines were established from samples of decidua from first trimester pregnancies. Apoptosis was assayed by flow cytometry. PRL production, as a marker of decidualization, was determined by enzyme-linked immunosorbent assay. MAIN RESULTS AND THE ROLE OF CHANCE: DSCs were resistant to a variety of apoptosis-inducing substances. Nevertheless, DSC underwent apoptosis during decidualization in culture, with cAMP being essential for both apoptosis and differentiation. In addition, culture supernatants from decidualized DSC induced apoptosis in undifferentiated DSC, although paradoxically these supernatants decreased the spontaneous apoptosis of decidual lymphocytes. Exogenously added PRL did not induce apoptosis in DSC and an antibody that neutralized the PRL receptor did not decrease the apoptosis induced by supernatants. LIMITATIONS, REASONS FOR CAUTIONS: Further studies are needed to examine the involvement of other soluble factors secreted by decidualized DSC in the induction of apoptosis. WIDER IMPLICATIONS OF THE FINDINGS: The present results indicate that apoptosis of DSC occurs in parallel to differentiation, in response to decidualization signals, with soluble factors secreted by decidualized DSC being responsible for triggering cell death. These studies are relevant in the understanding of how the regression of decidua, a crucial process for successful pregnancy, takes place. STUDY FUNDING/COMPETING INTERESTS: This work was supported by the Consejería de Economía, Innovación y Ciencia, Junta de Andalucía (Grant CTS-6183, Proyectos de Investigación de Excelencia 2010 to C.R.-R.) and the Instituto de Salud Carlos III, Ministerio de Economía y Competitividad, Spain (Grants PS09/00339 and PI12/01085 to E.G.O.). E.L.-D. was supported by fellowships from the Ministerio de Educación y Ciencia, Spain and the University of Granada. The authors have no conflict of interest.

Differential expression and regulation of Tdo2 during mouse decidualization.

Tryptophan 2,3-dioxygenase (Tdo2) is a rate-limiting enzyme which directs the conversion of tryptophan to kynurenine. The aim of this study was to examine the expression and regulation of Tdo2 in mouse uterus during decidualization. Tdo2 mRNA was mainly expressed in the decidua on days 6-8 of pregnancy. By real-time PCR, a high level of Tdo2 expression was observed in the uteri from days 6 to 8 of pregnancy, although Tdo2 expression was observed on days 1-8. Simultaneously, Tdo2 mRNA was also detected under in vivo and in vitro artificial decidualization. Estrogen, progesterone, and 8-bromoadenosine-cAMP could induce the expression of Tdo2 in the ovariectomized mouse uterus and uterine stromal cells. Tdo2 could regulate cell proliferation and stimulate the expression of decidual marker Dtprp in the uterine stromal cells and decidual cells. Overexpression of Tdo2 could upregulate the expression of Ahr, Cox2, and Vegf genes in uterine stromal cells, while Tdo2 inhibitor 680C91 could downregulate the expression of Cox2 and Vegf genes in uterine decidual cells. These data indicate that Tdo2 may play an important role during mouse decidualization and be regulated by estrogen, progesterone, and cAMP.

LGR4 expressed in uterine epithelium is necessary for uterine gland development and contributes to decidualization in mice.

In previous work we generated mice with a tissue specific ablation of a leucine-rich repeat containing G-protein-coupled receptor 4 (Lgr4) using the Keratin-5 (K5) Cre transgenic mouse strain (Lgr4(K5 KO)). Interestingly, the Lgr4(K5 KO) female mice were subfertile, and their embryos had impaired development. Notably, the contributions of uterine development to the subfertility phenotype were not elucidated in the previous report. In a readdress, the following study explores uterine aberration in Lgr4(K5 KO) female mice. Histological analysis revealed that the uteri of Lgr4(K5 KO) mice displayed altered epithelial differentiation characterized by a reduction in the number of uterine glands. Furthermore, Lgr4 deletion led to the reduced expression of morphoregulatory genes related to the Wnt signaling pathway. Additionally, the uteri of the Lgr4(K5 KO) mice lost the ability to undergo induced decidualization. Quantitative reverse transcription-polymerase chain reaction (qRT-PCR) analysis and administration of recombinant leukemia inhibitory factor (LIF) demonstrated that the impaired decidualization in Lgr4(K5 KO) mice resulted from the decreased secretion of LIF concurrent with a reduction in uterine gland count. Thus, we propose that LGR4 contributes to uterine gland development, which supports decidualization during pregnancy.